The Hela Cell Line and The HeLa Cell Culture

HeLa cells

HeLa cells stained with Hoechst 33258 (blue)
HeLa cells stained with Hoechst 33258 (blue)

HeLa cell culture protocol

HeLa cells are the most widely used cancer cell lines in the world. These cells were taken from a lady called Henrietta Lacks from her cancerous cervical tumour in 1951 which today is known as the HeLa cells. These were the very first cell lines to survive outside the human body and “grow”.

Some points to remember when culturing HeLa cells.

HeLa cells are adherent cells and hence stick to the bottom of the flask or the container used for growing the HeLa cells. It also has the ability to grow in suspension.

HeLa cells are best split or passaged when they are 70 to 80% confluent.

Use minimum of trypsin as they will increase the chances to develop trypsin-resistance cells.

As a rule cells should not be used up to a certain passage, it is thought that HeLa cells should be used up to a maximum of 20 passages.

Both DMEM and RPMI 1640 growth medium can be used to culture HeLa cells.

The doubling time for HeLa cells are 23-24 hours.

Media requirement for HeLa cells

Media (DMEM or RPM1) supplemented with 10% fetal calf serum, antibiotics (penicillin 100 units/ml + streptomycin 100 microgram/ml), 5% CO2 + 95% atmosphere at 37 degrees.

HeLa cells dividing over 27hrs cells round up grow hair then divide

HeLa Cells of Henrietta Lacks

HeLa Cells of Henrietta Lacks
HeLa Cells of Henrietta Lacks

HeLa Cells are the most widely used cell line in the world. This book looks at these immortal cell line in more detail.


Protocol for culturing HeLa cells:

Preheat the media, FCS, trypsin-EDTA at 37 degree in a water bath for a minimum of 30-45 minutes to ensure they are at 37 degrees.

Take the flask of HeLa cells out of the incubator and aspirate the old media out using a sterile pump.

Wash the adherent cells with new prewarmed media and aspirate to remove traces of the old media.

Add enough trypsin-EDTA to cover the cells (example: 5ml for a medium T75 flask) ands place the cells back in the incubator. Take a note of the time and wait. Take the container out after 2 minutes to see if detachment has taken place by viewing under the microscope.

After detachment, add complete media and FCS to the cells to deactivate the trypsin and pipette gently to disperse the cells.

Centrifuge the cells at 1000 rpm in a universal or a centrifuge tube depending on the volume.

Resuspend the cells in a small volume of complete media and count the cells using a haemocytometer or a cell counter.

Culture the cells in a new flask at a density of requirement and experimental need. Remember the doubling time of approximately 24 hours. Remember the split the cells very thinly over the weekends.

Dividing HeLa Cells


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